This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our laboratory has remained committed to delineating intrinsic cellular mechanisms associated with stem cell plasticity and differentiation. More specifically, for much of the last year we have focused on cellular mechanisms affected by Numb, an intracellular adaptor protein, which to date has been primarily characterized as an antagonist of the cell fate determinant, Notch, although it is not very clear how Numb mediates this function. Recently, the expression of two novel Numb isoforms, Numb 5 and Numb 6, was observed in neuroblastoma cells. Unlike the 4 prototypical Numb isoforms, Numbs 5 and 6 have been shown to potentiate Notch signaling rather than behave as antagonists. Numb 5 even recapitulates the effect of adding the Notch ligand, Delta, to neural stem cells (NSCs) by driving differentiation to gliogenesis instead of neurogenesis, the normal consequence of Notch antagonism. We are seeking to use Numb 5 and 6 as tools to dissect Numb function by exploiting the structural differences between these novel isoforms and the published Numb isoforms. We believe that a Proline rich region, which is missing in Numbs 5 and 6 possesses the structural information to allow Numb to be a Notch antagonist. Therefore, by using approaches designed to reveal novel direct protein-protein interactions during NSC differentiation, we hope to show that Numb 5 and 6 differ from the other Numb isoforms in terms of unique interactions with other factors associated with Notch antagonism and more broadly, neurogenesis. Already, we have been able to identify atypical Protein Kinase C zeta (aPKCzeta) as being in a complex with Numbs 1 and 2, but not the other isoforms. This observation is interesting because Numb and aPKC have both been shown to be components of cellular mechanisms associated with cell fate. Moreover, aPKC can be a regulator of Insulin signaling, another interest of this laboratory as it relates to cell fates of multi-lineage neural stem cells. In summary, we will continue to exploit the structural differences among the isoforms of Numb to learn how Numb modulates Notch signaling and more broadly, how Numb is involved in the regulation of NSC cell fate decisions.
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